We wish to continue our investigation into the mechanism of action of delta5-3-ketosteroid isomerase. This enzyme is a key enzyme in the biosynthesis of steroids, and a knowledge of its mechanism should allow selective modification by inhibitors, and consequently regulation of steroid biosynthesis. We will use our knowledge of the mechanism of delta5-3-ketosteroid isomerase to synthesize specific irreversible inhibitors of this enzyme which may be therapeutically useful in treating certain hormone-dependent cancers. We have already synthesized 8 active-site directed irreversible inhibitors of this enzyme. These fall into two main classes: 1) 3 Beta-oxiranes of dihydrotestosterone derivatives and 2) 17 Beta-oxiranes of estradiol and dehydroepiandrosterone derivatives. We are currently in the process of determining the site of attachment of one of these inhibitors, spiro-3beta-oxiranyl-5-androstan-17 beta-ol to get more detailed information about the active site. In addition to continuing this work, the site of attachment of a suitable 17 Beta-oxirane will be determined to determine if two modes are possible for binding of substrates (inhibitors) to this enzyme. This hypothesis will also be tested with other potential irreversible inhibitors. Kinetic studies will give information not previously obtainable about active site pKa's. These inhibitors, and other suitable derivatives which will be synthesized, will be tested for anti-cancer activity at both the National Cancer Institute and the University of Maryland Medical School.